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Clonal, self-renewing and differentiating human and porcine urothelial cells, a novel stem cell population.

Larsson HM, Gorostidi F, Hubbell JA, Barrandon Y, Frey P - PLoS ONE (2014)

Bottom Line: By examining an epithelial stem cell signature marker, we found p63 to correlate with the self-renewal capacity of the isolated human urothelial clonal populations.We isolated cells from porcine ureter, urethra and bladder that were clonogenic and capable of self-renewal and differentiation into fully mature urothelium.In conclusion, we could isolate human and porcine cell populations, behaving as urothelial stem cells and showing clonogenicity, self-renewal and, once re-implanted, morphological differentiation.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Regenerative Medicine and Pharmacobiology, Institute for Bioengineering, School of Life Sciences and School of Engineering, Ecole Polytechnique Fédérale de Lausanne, Lausanne, Switzerland.

ABSTRACT
Although urothelial progenitor-like cells have been described in the human urinary tract, the existence of stem cells remains to be proven. Using a culture system that favors clonogenic epithelial cell growth, we evaluated and characterized clonal human urothelial cells. We isolated human urothelial cells that were clonogenic, capable of self-renewal and could develop into fully differentiated urothelium once re-implanted into the subcapsular space of nude mice. In addition to final urothelial cell differentiation, spontaneous formation of bladder-like microstructures was observed. By examining an epithelial stem cell signature marker, we found p63 to correlate with the self-renewal capacity of the isolated human urothelial clonal populations. Since a clinically relevant, long-term model for functional reconstitution of human cells does not exist, we sought to establish a culture method for porcine urothelial cells in a clinically relevant porcine model. We isolated cells from porcine ureter, urethra and bladder that were clonogenic and capable of self-renewal and differentiation into fully mature urothelium. In conclusion, we could isolate human and porcine cell populations, behaving as urothelial stem cells and showing clonogenicity, self-renewal and, once re-implanted, morphological differentiation.

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Porcine clonal urothelial cells arising from a single bladder cell.(A, B and C) Porcine urethelial holoclone, meroclone and paraclone cultures arising from a single bladder cell. (D and E) Growth curves of a porcine urothelial holoclone. (F) In vivo urothelial differentiation of porcine bladder urothelial holoclone pellets implanted into the subcapsular space of the Swiss nu/nu mice, expressing cytokeratin 7, uroplakin-2 and uroplakin 3 (scale bars, 10 µm). Note the “micro-bladder” like structure.
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pone-0090006-g005: Porcine clonal urothelial cells arising from a single bladder cell.(A, B and C) Porcine urethelial holoclone, meroclone and paraclone cultures arising from a single bladder cell. (D and E) Growth curves of a porcine urothelial holoclone. (F) In vivo urothelial differentiation of porcine bladder urothelial holoclone pellets implanted into the subcapsular space of the Swiss nu/nu mice, expressing cytokeratin 7, uroplakin-2 and uroplakin 3 (scale bars, 10 µm). Note the “micro-bladder” like structure.

Mentions: Studying the in vitro behavior of cells harvested from the porcine bladder, ureter and urethra, we observed the same clonogenic capacity as the cells harvested from the human ureter. Porcine clonogenic urothelial cells were, as for the human cells, classified as urothelial holoclones, meroclones or paraclones, depending on the growth capacities. (Figure 4A–C, 5A–C and 6A–C) (Table 2, 3 and 4). Compared to human holoclones, porcine holoclones had a much greater growth capacity. It could be demonstrated that after 38 days of passaging, porcine urothelial holoclone showed 30 population doublings and thereafter were still in a growing phase (Figure 4D–E, 5D–E and 6D–E).


Clonal, self-renewing and differentiating human and porcine urothelial cells, a novel stem cell population.

Larsson HM, Gorostidi F, Hubbell JA, Barrandon Y, Frey P - PLoS ONE (2014)

Porcine clonal urothelial cells arising from a single bladder cell.(A, B and C) Porcine urethelial holoclone, meroclone and paraclone cultures arising from a single bladder cell. (D and E) Growth curves of a porcine urothelial holoclone. (F) In vivo urothelial differentiation of porcine bladder urothelial holoclone pellets implanted into the subcapsular space of the Swiss nu/nu mice, expressing cytokeratin 7, uroplakin-2 and uroplakin 3 (scale bars, 10 µm). Note the “micro-bladder” like structure.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3935977&req=5

pone-0090006-g005: Porcine clonal urothelial cells arising from a single bladder cell.(A, B and C) Porcine urethelial holoclone, meroclone and paraclone cultures arising from a single bladder cell. (D and E) Growth curves of a porcine urothelial holoclone. (F) In vivo urothelial differentiation of porcine bladder urothelial holoclone pellets implanted into the subcapsular space of the Swiss nu/nu mice, expressing cytokeratin 7, uroplakin-2 and uroplakin 3 (scale bars, 10 µm). Note the “micro-bladder” like structure.
Mentions: Studying the in vitro behavior of cells harvested from the porcine bladder, ureter and urethra, we observed the same clonogenic capacity as the cells harvested from the human ureter. Porcine clonogenic urothelial cells were, as for the human cells, classified as urothelial holoclones, meroclones or paraclones, depending on the growth capacities. (Figure 4A–C, 5A–C and 6A–C) (Table 2, 3 and 4). Compared to human holoclones, porcine holoclones had a much greater growth capacity. It could be demonstrated that after 38 days of passaging, porcine urothelial holoclone showed 30 population doublings and thereafter were still in a growing phase (Figure 4D–E, 5D–E and 6D–E).

Bottom Line: By examining an epithelial stem cell signature marker, we found p63 to correlate with the self-renewal capacity of the isolated human urothelial clonal populations.We isolated cells from porcine ureter, urethra and bladder that were clonogenic and capable of self-renewal and differentiation into fully mature urothelium.In conclusion, we could isolate human and porcine cell populations, behaving as urothelial stem cells and showing clonogenicity, self-renewal and, once re-implanted, morphological differentiation.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Regenerative Medicine and Pharmacobiology, Institute for Bioengineering, School of Life Sciences and School of Engineering, Ecole Polytechnique Fédérale de Lausanne, Lausanne, Switzerland.

ABSTRACT
Although urothelial progenitor-like cells have been described in the human urinary tract, the existence of stem cells remains to be proven. Using a culture system that favors clonogenic epithelial cell growth, we evaluated and characterized clonal human urothelial cells. We isolated human urothelial cells that were clonogenic, capable of self-renewal and could develop into fully differentiated urothelium once re-implanted into the subcapsular space of nude mice. In addition to final urothelial cell differentiation, spontaneous formation of bladder-like microstructures was observed. By examining an epithelial stem cell signature marker, we found p63 to correlate with the self-renewal capacity of the isolated human urothelial clonal populations. Since a clinically relevant, long-term model for functional reconstitution of human cells does not exist, we sought to establish a culture method for porcine urothelial cells in a clinically relevant porcine model. We isolated cells from porcine ureter, urethra and bladder that were clonogenic and capable of self-renewal and differentiation into fully mature urothelium. In conclusion, we could isolate human and porcine cell populations, behaving as urothelial stem cells and showing clonogenicity, self-renewal and, once re-implanted, morphological differentiation.

Show MeSH